Ricin is a plant toxin produced by the castor bean (Ricinus communis) that is extremely toxic to mammalian cells and is classified as a select agent by the US Department of Health and Human Services (HHS) and a category B priority pathogen by the Center for Disease Control (CDC). Although there have been attempts to generate an effective vaccine against ricin, no recombinant vaccine has yet been approved for human use and there are no effective treatment measures against ricin exposure. Therefore there is an urgent need for therapeutics to protect ricin-exposed individuals. To gain understanding into the mechanism of ricin induced cell death, we will take advantage of the unique experimental features of C. elegans that have made it a leading model organism in nearly all areas of biological research. Our primary objective in this proposal is to establish C. elegans as a new whole animal model to study ricin intoxication and to translate our findings to mammals. In preliminary studies, we fed C. elegans with either E. coli expressing ricin A-chain (RTA) or recombinant RTA purified from E. coli and demonstrated that RTA is lethal to C. elegans at extremely low doses. In agreement with mammalian data, we showed that ricin uses a clathrin and a raft independent endocytosis pathway to enter the C. elegans intestinal epithelial cells and entry to the early endosome is critical for cytotoxicity. We identified a novel role for the two major phagocytosis/engulfment pathways in mediating ricin sensitivity in the C. elegans intestine. These results provided support for the development of C. elegans as a new multicellular model for molecular genetic dissection of the mechanisms of ricin transport and cell death. We will examine transport of ricin in the worm intestine, a classic polarized epithelium, in the context of a living animal. C. elegans mutants defective in intracellular trafficking pathways will be used to validate C. elegans as a relevant model for mechanistic analysis of ricin uptake and transport and to identify critical steps in ricin transport that lead to cytotoxicity. We will exploit powerful features of C. elegans genetics to isolate new mutants defective in ricin intoxication. The proposed studies will lead to elucidation of the molecular details of ricin transport and will lay the foundation for identification of inhibitors that can prevent ricin induced cell death. PUBLIC HEALTH RELEVANCE: Our primary objective in this application is to establish C. elegans as a new model to study the mechanism of ricin intoxication and to translate our findings to humans. These studies are significant because currently there are no vaccines, antidotes or any other treatment measures against ricin exposure. These studies will lay the foundation for identification of inhibitors that can protect ricin exposed individuals.